Many cell types must become polarized to function properly within a tissue or to generate distinct daughter cells. A quintessential example of this phenomenon is observed in the eggs of Caenorhabditis elegans. Shortly after fertilization, the C. elegans zygote undergoes a dramatic, stereotyped polarization that segregates polarity factors and cell fate determinants to the anterior and posterior ends of the egg. These asymmetries are essential for cellular differentiation and organismal development. Genetic analyses have demonstrated an essential role for the conserved Serine/Threonine kinase PAR-1 in this process. Specifically, loss of par-1 protein activity in the zygote results in symmetric distributin of normally asymmetric factors. In this proposal I will combine new technologies with classical techniques to analyze PAR-1 function in live embryos. In Aim1, I will use optogenetic and Cas9 genome editing technologies to assign function to the different PAR-1 domains and test the role of PAR-1 asymmetry in patterning the zygote. In Aim 2, I will conduct an unbiased chemical genetic screen to identify critical PAR-1 substrates. This work will uncover the molecular mechanisms used by PAR-1 to pattern the zygote and regulate cell and organismal polarity.